Muscle contraction in all animals is switched on and off by changes in the physiological level of free calcium in the sarcoplasm. Regulatory proteins, which bind Ca ions and confer a Ca ions dependency to actomyosin interactions with ATP are located on either the thin filaments, the myosin, or in most cases on both. Thin filament-linked regulation involves an interaction of Ca ions and the troponin-tropomyosin complex, modifying the activity of actin; in myosin regulation, the myosin molecule is directly modulated, and this presumably results from a Ca ions -dependent interaction of the light and heavy chains of myosin. In vertebrates, skeletal muscle possesses a thin filament-linked system, and smooth muscles are myosin-regulated. In our comparative survey, we have found the dual system of regulation in most invertebrates. We also have obtained suggestive evidence of possible dual-regulation in vertebrate skeletal muscle, but more direct and conclusive evidence is required. Our studies therefore will be extended to determine if vertebrate muscles are also dual-regulated. Vascular and visceral smooth muscles, cardiac, and both fast and slow skeletal muscles will be investigated. The sites of Ca ions regulation will be determined by well established ATPase assays and the Ca ions binding properties of the regulatory proteins studied.